Characterization of genetic diversity in Curcuma longa L. cultivars using Start Codon Targeted (SCoT) markers

Abstract

Curcuma longa L. (Family- Zingiberaceae), commonly called turmeric, is one of the most important commercial spices. Being a center of turmeric's diversity, India boasts of more than fifty cultivars with variable agronomic characters and curcumin content. However, the quest for calibrating their genetic variability remains debatable as popular molecular marker-based studies have yielded inconsistent results. A promising PCR-based approach, known as the Start Codon Targeted (SCoT)-PCR system has been established to be more efficient than random DNA marker systems because this technique can generate more information on genetic polymorphism patterns. Hence, the present study proposes to use Start Codon Targeted (SCoT) markers to assess inter-cultivar genetic diversity of turmeric. We report an orthogonal L16 (45) design-based approach to investigate the influence of five crucial factors (dNTPs, Taq DNA polymerase, primer, template DNA and annealing temperature of primer) on the efficiency of the polymerase chain reaction (PCR) for development of an optimized Start Codon Targeted (SCoT) amplification protocol using twenty-seven different cultivars of C. longa. The optimized PCR protocol yields best results with a 20 uL reaction mixture, containing 10×PCR Buffer (with 15 mM MgCl2), 2.5 mM dNTPs, 10 mM SCoT primer, Taq polymerase (2 U), template DNA (50 ng), which was verified by using twenty-five SCoT primers. This standardized rapid protocol of SCoT-PCR reported here was employed to examine genetic diversity among twenty-seven Indian cultivars of turmeric, revealing high range of polymorphism ranging from 81.25 % to 100 %. Unweighted pair group method of arithmetic averages (UPGMA) cluster analysis of the SCoT marker data segregated the twenty-seven cultivars into two main clusters. The first cluster comprised of nine cultivars (36 % of the cultivars) which was further divided into three sub-clusters and the second one consisted of eighteen cultivars (64 %) which further divided into four sub-clusters. From the results of this study, it can be inferred that the employment of SCoT markers can be an effective tool for analysing genetic variability, phylogenetic relationships and develop a rich genetic resource management system for C. longa cultivars.